电子自旋共振经典讲座演示文稿

电子自旋共振经典讲座演示文稿当前第1页\共有23页\编于星期五\20点优选电子自旋共振经典讲座当前第2页\共有23页\编于星期五\20点FreeRadicalsandEPRMoleculeswithoneormoreunpairedelectronQuantummechanics:unpairedelectronshavespinandchargeandhencemagneticmomentElectronicspincanbeineitheroftwodirections(formallyupordown)ThetwospinstatesundernormalconditionsareenergeticallydegenerateEnergeticdegeneracylostwhenexposedtoanexternalmagneticfield当前第3页\共有23页\编于星期五\20点TheEPRexperimentPutsampleintoexperimentalmagneticfield(B)Irradiate(microwavefrequencies)Measureabsorbanceofradiationasf(B)Weil,Bolton,andWertz,1994,“ElectronParamagneticResonance”当前第4页\共有23页\编于星期五\20点TheEPRspectrometerElectromagnetMicrowavesourceanddetector(typicallyXband,~9.5GHz)Modulationofmagneticfieldandphase-sensitivedetectionSpectrum1stderivativeWeil,Bolton,andWertz,1994,“ElectronParamagneticResonance”当前第5页\共有23页\编于星期五\20点TheEPRspectrumA1stderivativespectrumisobtainedfromtheunpairedelectronhn=gBb0gisacharacteristicofthechemicalenvironmentoftheunpairedelectron;forfreeradicalsitisnear2.00;canvarywidelyfortransitionmetalcentersComplicated/enhancedbyhyperfineinteractionswithnucleiwithnon-zerospin当前第6页\共有23页\编于星期五\20点ThehyperfineeffectThemagneticfieldexperiencedbytheunpairedelectronisaffectedbynearbynucleiwithnon-zeronuclearspinWeil,Bolton,andWertz,1994,“ElectronParamagneticResonance”,NewYork:WileyInterscience.当前第7页\共有23页\编于星期五\20点HyperfinesplittingofEPRspectraThemagnitudeofthesplittingandthenumberoflinesdependupon:Thenuclearspinoftheinteractingnucleus#oflines=2n(I+½)soI=½gives2lines,etc.ThenucleargyromagneticratioThemagnitudeoftheinteractionbetweentheelectronicspinandthenuclearspinMagnitudeofthesplittingtypicallydecreasesgreatlywithincreasingnumbersofbondsbetweenthenucleusandunpairedelectron当前第8页\共有23页\编于星期五\20点当前第9页\共有23页\编于星期五\20点Hyperfinesplittingsareadditive当前第10页\共有23页\编于星期五\20点DirectEPRanalysisofaradicalRadicalcannotbediatomicRadicalmustbeavailableatadetectableconcentrationAtleastmetastableFrozensolutiontogreatlydecreaseradicaldecayCangreatlycomplicatethespectrumduetoanisotropyContinuousformationinsideresonatorEnzymaticradicalformationFlowexperimentRadicalcharacterizedbyhyperfineanalysis当前第11页\共有23页\编于星期五\20点DirectEPRofatyrosylradicalGunther,M.R.,Sturgeon,B.E.,andMason,R.P.,FreeRadic.Biol.Med.28:709-719,2000当前第12页\共有23页\编于星期五\20点Spintrapping:whendirectEPRisnotconvenientorpossibleUnstablefreeradicalreactswithdiamagneticmolecule(thespintrap)toformarelativelystablefreeradicalThevastmajorityofspintrapsformradicaladductsthroughtheadditionoftheradicaltothetraptoformanitroxideradical2majorclassesoftraps:nitronesandnitrosocompounds当前第13页\共有23页\编于星期五\20点AdvantagesofthenitronesReactwithavarietyofdifferentfreeradicalstoformnitroxideadductsRC.,RO.,RS.,insomecasesRN.AdductsareoftenquitestableNotterriblytoxicsoamenabletoinvivo/exvivospintrapping当前第14页\共有23页\编于星期五\20点NitronespintrapsDMPO,5,5-dimethylpyrrolineN-oxidePBN/4-POBN,phenyl-N-t-butylnitrone当前第15页\共有23页\编于星期五\20点EPRspectrafromDMPOadducts当前第16页\共有23页\编于星期五\20点EPRspectrafrom4-POBNadducts当前第17页\共有23页\编于星期五\20点NitrosospintrapsFreeradicaladdstothenitrogenatomofaC-nitrosocompound2-methyl-2-nitrosopropane,MNP3,5-dibromo-4-nitrosobenzenesulfonate当前第18页\共有23页\编于星期五\20点EPRspectrafrommethylradicaladductsofnitrosotraps当前第19页\共有23页\编于星期五\20点DMPO-trappingthetyrosylradicalOxidizetyrosinewithHRP/H2O2Gunther,M.R.,etal.,Biochem.J.330:1293-1299,1998.当前第20页\共有23页\编于星期五\20点Spintrap-derivedhyperfinefromMNPandMNP-d9EachlineintheEPRspectrafromMNPadductsisbroadenedbyhyperfinefromthe9equivalentprotonsonthespintrap当前第21页\共有23页\编于星期五\20点MNP-trappingthetyrosylradicalGunther,M.R.,etal.,Biochem.J.330:1293-1299,1998.当前第22页\共有23页\编于星期五\20点Whynotspintrap?Nitronespintraps,especiallyDMPOAdductscanintercon